Our observations in gnotobiotic mice and unrelated adult obese and lean humans indicate that there is a dynamic linkage between adiposity and gut microbial ecology. Project 1will test the hypothesis that thereis an identifiable shared set of organismal and gene lineages evident in the microbiota and microbiomes of obese compared to lean adult individuals, and that these obesity-associated organismal genes have predicted properties that could contribute to increased efficiency of energy harvest from a diet. Aim 1 -We will perform comparative metagenomic (DNA-based) sequence analysis of fecal samples obtained from young adult (25-35year old), vaginally delivered female MZtwin pairs who are obese (BMI^35 kg/m2), and MZ twin pairs who are lean (BMI 18.5-25kg/m2): five of the MZ twin pairs in each category (obese or lean) will be of European ancestry (total of 10 twin pairs). Five more twin pairs in each category will be African- American. Mothers of twin pairs will be used as reference controls since there is evidence for vertical transmission of the gut microbiota from a mother to her offspring. Two fecal samples will initially be collected per individual over a one-month interval. Each fecal community DNA sample from each patient will be subjected to 16S rRNA sequence-based enumeration using 'universal'bacterial, and archaeal primers. The two DNA samples from each individual will be pooled, and -40 Mb of DNA sequence will be generated using the GS20 pyrosequencer. We will compare communities using the UniFrac metric developed by the Knight lab, and employ existing algorithms, developed from our studies of obese and lean mice, to compare their microbiomes. The collection of both short-term (1-month) and longer-term followup fecal samples (at 12, 24, 36, and 48 months) from MZ twin pairs and their mothers will allow us to examine the structureof their microbiotas and microbiomes over time relative to co-twin, mother, and BMI.To determine whether genetic background of the host versus a shared mother plays a dominant role in selecting a microbiota and its microbiome, we will perform a comparable analysis of the microbiomes of obese and lean 25-35year-old female European ancestry and African-American dizygotic twin pairs and their mothers. Aim 2 - The representation of KEGG and COG functional groups found in the fecal microbiomes of obese vs. lean MZ twin pairs will be correlated with fecal microbial community transcriptomes and metabolomes. cDNA libraries prepared from RNAs isolated from the same fecal samples previously used for whole microbiome shotgun DNA sequencing, will be characterized. Genes and transcripts identified in the fecal microbiome/transcriptome of obese vs. lean individuals will be placed onto KEGG metabolic pathways. The